In mammals, there are two biochemical pathways for triacylglycerol synthesis: the monoacylglycerol pathway, which happens exclusively in the small intestine (R. Lehner and A. Kuksis, Prog. Lipid Res., (1996) 35:169-201), and the glycerol-3-phosphate pathway, which takes place ubiquitously but most notably in the liver and in adipose tissue (R. M. Bell and R. A. Coleman, Annu. Rev. Biochem., (1980) 49:459-487). The monoacylglycerol pathway initiates from acyl coenzyme A:monoacylglycerol acyltransferase (MGAT) (EC 2.3.1.22). Within minutes of its appearance from the digestion of dietary fat in the lumen of the small intestine, 2-monoacylglycerol is acylated by MGAT to form diacylglycerol. Diacylglycerol is further acylated by acyl coenzyme A:diacylglycerol acyltransferase (DOGAT) (EC 2.3.1.20) to re-synthesize triacylglycerol, which is packaged into chylomicron lipoprotein particles that eventually secreted into the lymph. In the glycerol-3-phosphate pathway, two fatty acyl coenzyme A molecules are added to glycerol-3-phosphate to form phosphatidate. These reactions are followed by the removal of the phosphate group by phosphatidate phosphohydrolase to generate diacylglycerol. Diacylglycerol is then further acylated by DGAT to form triacylglycerol. Collectively, DGAT lies at the final step of both triacylglycerol synthesis pathways.
Two DGAT enzymes have been identified, which are designated as DGAT1 and DGAT2 (S. Cases, S. J. Smith, Y. W. Zheng, H. M. Myers, S. R. Lear, E. Sande, S. Novak, C. Collins, C. B. Welch, A. J. Lusis, S. K. Erickson and R. V. Farese, Jr, Proc. Natl. Acad. Sci. USA, (1998) 95:13018-13023; P. Oelkers, A. Behari, D. Cromley, J. T. Billheimer and S. L. Sturley, J. Biol. Chem., (1998) 273:26765-26771; S. Cases, S. J. Stone, P. Zhou, E. Yen, B. Tow, K. D. Lardizabal, T. Voelker and R. V. Farese, Jr., J. Biol. Chem., (2001) 276:38870-38876). Although they carry out identical enzymatic reactions, DGAT1 and 2 are encoded by two different genes that bear little sequence homology. Functionally, these two enzymes might have different physiological importance in vivo. DGAT1 knockout mice exhibit resistance towards the challenge of a high fat diet to become obese (S. J. Smith, S. Cases, D. R. Jensen, H. C. Chen, E. Sande, B. Tow, D. A. Sanan, J. Raber, R. H. Eckel and R. V. Farese, Jr., Nat. Genet., (2000) 25:87-90). They are physically more active, possess a higher metabolic rate (H. C. Chen and R. V. Farese, Jr., Trends Cardiovasc. Med., (2000) 10:188-192) and appear to have a greater insulin sensitive (H. C. Chen, S. J. Smith, Z. Ladha, D. R. Jensen, L. D. Ferreira, L. K. Pulawa, J. G. McGuire, R. E. Pitas, R. H. Eckel and R. V. Farese, Jr., J. Clin. Invest., (2002) 109:1049-1055). In contrast, DGAT2 knockout mice exhibit phenotypes such as lipopenia and skin barrier abnormalities, resulting in death soon after birth (S. J. Stone, H. M. Myers, S. M. Watkins, B. E. Brown, K. R. Feingold, P. M. Elias and R. V. Farese, Jr., J. Biol. Chem., (2004) 279:11767-11776).
DGAT1 belongs to the acyl coenzyme A:cholesterol acyltransferase (ACAT, EC 2.3.1.26) gene family. ACAT is the enzyme responsible for synthesis of cholesteryl ester using cholesterol and long-chain fatty acyl-coenzyme A as substrates. ACAT and DGAT have considerable biochemical similarities: both enzymes are proteins with multiple transmembrane domains and reside in the endoplasmic reticulum (ER). Both catalyze the reaction involving the transfer of an acyl-moiety of acyl coenzyme A to a hydrophobic substrate.
Both DGAT and ACAT have been considered as potential therapeutic targets. For DGAT, it has been proposed that inhibiting the synthesis of triacylglycerol would benefit in reducing weight, improve insulin sensitivity and reduce hepatic and circulating lipid content. For ACAT, it had been considered as a therapeutic target for cholesterol lowering and for anti-atherosclerosis.
During the past 20 years, numerous ACAT inhibitors have been developed (D. R. Sliskovic, J. A. Picard, B. R. Krause, “ACAT inhibitors: the search for a novel and effective treatment of hypercholesterolemia and atherosclerosis”, Prog. Med. Chem., (2002) 39:121-171). It is conceivable that due to the homology between ACAT and DGAT1, some ACAT inhibitors might possess DGAT inhibitory activities. Among them, thianecarboxamids are a series potent inhibitors that inhibits both hepatic and macrophage ACAT effectively (U.S. Pat. No. 5,491,152; R. G. Wilde, J. T. Billheimer, S. J. Germain, E. A. Hausner, P. C. Meunier, D. A. Munzer, J. K. Stoltenborg, P. J. Gillies, D. L. Burcham, S. M. Huang, J. D. Klaczkiewicz, S. S. Ko, R. R. Wexler, “ACAT inhibitors derived from hetero-Diels-Alder cycloadducts of thioaldehydes”, Bioorg. Med. Chem., (1996) 4:1493-1513). In the current invention, we provide evidence that this chemical series indeed possess potent inhibitory activities against DGAT.